The mechanisms controlling T helper differentiation into the Th1 and Th2 lineages are not fully understood. These studies investigate the role of the SRC-family kinase Lck in Th2 differentiation. Th2 cells secrete anti-inflammatory cytokines IL-4, IL-5, IL-10, and IL-13 and mediate protection against extracellular pathogens. Th1 cells are known to produce pro-inflammatory cytokines IFN-y and TNF-a and mediate protection against intracellular pathogens. The balance between the Th1 and Th2 lineages must be maintained, because improper development of TM or Th2 effector lineages can lead to disease. Th2 cytokines are grouped into two genetic loci. The cytokines IL-5, IL-13, and IL-4 are linked in what is referred to as the Th2 cytokine locus, while IL-10 is found elsewhere in the genome. GATA-3 controls the expression of these cytokines, although the mechanism differs depending on the locus. Past studies have suggested that Lck plays a role in Th2 differentiation and/or function, but these studies did not determine the mechanism by which Lck functions. Lck deficient Th2 skewed cells have little to no expression of the Th2 cytokine IL-4, normal IL-10 expression, and synthesize the Th1 cytokine IFN-y. Preliminary studies indicate that Lck deficient Th2 skewed cells inappropriately express Th1 specific transcription factor T-bet, and have reduced expression of Th2 specific transcription factor GATA-3. Because IL-10 production is normal in the mutants, the defect may be specific for the Th2 cytokine locus. We hypothesize that Lck mediates full Th2 lineage commitment via maintenance of GATA-3 expression, and in the absence of Lck, GATA-3 is inhibited through ectopic expression of T-bet, resulting in a defect in Th2 effector functions due to defective expression of cytokines found at theTh2 cytokine locus. These studies will focus on two areas. The first aim will determine if Lck is required for expression of all cytokines associated with the Th2 cytokine locus or only IL-4. The second aim will determine if ectopic T-bet inhibits GATA-3 expression in the absence of Lck. T-bet has been shown to inhibit GATA-3 expression and function when overexpressed. These studies will provide insight into the mechanisms that control T helper differentiation. PUBLIC HEALTH RELEVANCE: The rates of Th2 mediated disorders, such as asthma and allergy, have increased rapidly in past decades. A greater understanding of the factors that control Th1 and Th2 differentiation, such as Lck, is needed. These studies will help elucidate the mechanisms that mediate Th2 differentiation and potentially lead to treatments of Th2 mediate diseases, such as allergy and asthma. [unreadable] [unreadable] [unreadable]